Undifferentiated hESC/hiPSC cells were sorted for hTRA-1-81+ to control for the homogeneity of cells before RNA extraction. The quality and quantity of total input mRNA was determined on an Agilent BioAnalyzer 2100 using Agilent RNA 6000 Nano kit. One microgram of total RNA from each sample was then used as input for library preparation using Illumina TruSeq RNA Sample Prep Kit, following manufacturer’s instructions. Each paired-end library was prepared with an adaptor with unique index sequence. The size profile and quantity of resulting libraries were than determined on the BioAnalyzer 2100 with Agilent High Sensitivity DNA kit. These libraries were then pooled together at equal molar concentration and sequenced on an Illumina HiSeq 2000. All hESC and hiPSC samples for RNA-Seq analysis were prepared on the same day by the same person, and then sequenced simultaneously on the same run (except for hiPSC lines 1, 2 and 3; this did not affect the clustering). All fibroblasts samples were prepared and sequenced in the same manner as the pluripotent samples but on different days.
